We will establish simple and convenient methods to knock out specific gene expression in animals. Antisense oligonucleotides (ASO) and antisense genes (ASG) can selectively suppress gene function by interacting with the message of interest to eliminate a specific protein. This approach is currently unfeasible in animals because of pharmacokinetic and transport (both into the cytoplasm and into the nucleus) considerations. We will develop, characterize and use in vivo, high efficiency delivery systems for ASO and ASG. Two delivery systems will be used: 1) previously described liposomes that deliver their contents into the cytoplasm of cells, 2) a novel approach which directly attaches targeting ligands to plasmid DNA, in effect constructing the delivery system on the DNA. For the second approach we have synthesized bis-acridine intercalators that contain nuclear localization peptide sequences (NLS) or ligands for surface receptors. The affinity of the intercalator-ligands for DNA and the avidity of the ligand-modified DNA for the target receptor as well as its extent of internalization will be measured on target cells. Distribution of radiolabeled targeted plasmids will be measured following i.v. administration in mice. Since transfection frequency is limited by nuclear entry of DNA, NLS-acridine conjugates will be attached to DNA to increase nuclear uptake. Factors that control targeting of the modified DNA into the nucleus will be studied in isolated nucleii and following microinjection of the complex into cells. The influence of increased nuclear localization of the plasmid on gene expression will be quantitated.